Toxic Effects Of Graphene Nanomaterials On Earthworms

Graphene is one of the most popular research topics in carbon nanomaterials that widely used in many fields due to its excellent physical and chemical properties.As the production and application amount is increasing,graphene will be inevitably released to the soil environment,resulting a potential ecological risk to soil organisms.In order to further understand the potential risks of graphene nanomaterials to soil organisms,the acute toxic effects of graphene nanomaterials on soil model organism earthworm(Eisenia fetida)were investigated.The results are summarized as follows:1.In the acute toxicity tests(artificial soil assay),different concentrations of graphene(0,100,300,1000 and 3000 mg·kg-1)had no significant effects on the mortality and reproduction rate of earthworms after exposure for 7,14 and 28 days.After acute exposure 7 and 14 d,although different concentrations of graphene treatment had little effect on the growth of adult earthworms,the growth of juvenile earthworms was significantly inhibited in the graphene treatment group at high concentrations(3000 mg·kg-1).Avoidant behavior experiments showed that adult earthworms and juvenile earthworms did not achieve obvious avoidance behavior(<80%)after exposure to different concentrations of graphene for 48 h,but with the increase of graphene concentration,the avoidance behavior of earthworms increased.2.There were no significant changes in soluble protein and catalase(CAT)in earthworms after 7 d treatment with different concentrations of graphene.However,superoxide dismutase(SOD)in high concentration(3000 mg·kg-1)group was significantly inhibited(P<0.05).After 14 d treatment with different concentrations of graphene,the soluble protein,CAT and SOD in earthworms were significantly decreased with the increase of graphene concentration(P<0.05),indicating that graphene could interfere with the antioxidant defense system in earthworms with the increase of graphene concentration and the prolongation of exposure time.3.We used matrix assisted laser desorption and ionization mass spectrometry imaging(MALDI-MSI)-based untargeted metabolomics to investigate the metabolic response of juvenile earthworms to graphene exposure in soil tests.The results reveal that different concentrations of graphene exposure significantly disturbs earthworm metabolome,and graphene toxicity on earthworm shows non-concentration-dependent effect.Twelve metabolites were successfully screened as significantly change compounds in earthworms for the exposure of graphene,including 6 amino acids(alanine,phenylalanine,proline,glutamate,arginine,and histidine),4 metabolites related to energy metabolism(maltose,glucose,malate,and succinate),1 compound involved in osmotic equilibrium(myo-inositol)and 1 molecule belongs to polyamines(spermidine).Most importantly,MALDI-MSI clearly and intuitively showed the changes in the distribution of 12 metabolites in earthworms.This work greatly improves our understanding of graphene nanoparticle toxicity to juvenile earthworms.Our data provided clear evidence that graphene had no effect on earthworm mortality and reproduction,but it inhibited the growth of juvenile earthworms,increased avoidance behavior,inhibited antioxidant enzyme activity,interfered with the normal metabolism of earthworm,and caused changes in the spatial distribution of metabolites,which provided a theoretical basis for the ecological risk assessment of graphene-contaminated soil.